Open access peer-reviewed chapter
Demographic information of 12 participants.
Abstract
This study is aimed at exploring if “naloxone” is detected in urine and water samples by dipping buprenorphine/naloxone film directly into these specimens. This study utilized 12 urine samples from 12 healthy participants who were not taking any medications with four samples added as a control. Sublingual generic buprenorphine/naloxone (8 mg/2 mg) film was dipped directly into these samples. They were sent to the ARUP laboratory for gas chromatography-mass spectrometry (GC/MS) quantitative analysis. The results were analyzed using IBM SPSS Statistics software. The results showed that “naloxone” was detected at high levels both in urine samples and in water, into which buprenorphine/naloxone film was dipped. In addition, the “naloxone” level was associated with the area of the film and the time in contact with the urine or water samples, but it was not affected by the urine concentration or the temperature of the specimens. This information will be useful for clinicians in identifying urine manipulation and interpreting urine drug test results and can help them for accurate monitoring of their patients’ treatment progress in opioid use disorder (OUD) treatment programs.
Keywords
- opioid
- urine
- buprenorphine
- norbuprenorphine
- creatinine
1. Introduction
Naloxone1 is an opioid antagonist that binds to
Naloxone has approximately 30–45 minute duration of action with a short half-life of 1.87–5.45 hours and reaches peak activity within 0.750–1.13 hours when taken sublingually [2, 3, 4]. It dissociates rapidly from opioid receptors within 6.5 minutes and has a low bioavailability of an estimated 3% through the sublingual route of administration with extensive first-pass metabolism [5]. Naloxone can be administered through various routes, which include intravenous, intramuscular, subcutaneous, endotracheal, sublingual, intralingual, submental and intranasal routes [6]. Naloxone, however, is estimated to be 50–250 times more potent when intravenously injected than orally administered [7]. Given this unique characteristic, naloxone is administered parenterally in order to increase its bioavailability, especially when it is used to reverse opioid effect in opioid overdose cases [8].
The effect of naloxone in combined buprenorphine/naloxone medication when taken sublingually is assumed to be clinically insignificant due to its poor absorption (<10%) and extensive first pass metabolism [4]. Therefore, combining buprenorphine and naloxone is strategized to deter unintended use such as intravenous injection and intranasal insufflation [9]. When taken through these routes, the addition of naloxone to buprenorphine antagonizes
While it is widely accepted that naloxone has a poor oral and sublingual bioavailability, many studies found “naloxone” detected at various levels in urine samples of patients who are on sublingual buprenorphine/naloxone medication [11, 12]. For example, Strickland and Burson found that 92.7% of urine samples from those who were on buprenorphine/naloxone medication had >30 ng/mL of “naloxone” detected [13]. At high levels within the system, naloxone may exert its antagonist effect on
In addition, many means of urine adulteration among the OUD patients who are on buprenorphine have been reported. One of the common methods of urine adulteration is “dipping” or “spiking” in which patients attempt to dissolve buprenorphine/naloxone film into urine samples directly. Heikman et al. reported that stable patients who are on buprenorphine/naloxone treatment had a median urine “naloxone” concentration of 60 ng/mL while that of unstable patients had a similar median of 70 ng/mL. The same study also found that the urine “naloxone” concentration in unstable patients ranged between 10 and 1700 ng/mL while that of the stable patients had a narrower window between 5 and 200 ng/mL [15]. This contrasting comparison of urine “naloxone” concentrations between stable and unstable patients raises the question of appropriate use of buprenorphine/naloxone therapy and the possibility of high “naloxone” levels in urine as an indication for urine adulteration.
Urine testing strategies have been developed to address this practice. Many previous studies discussed the use of elevated ratio between “buprenorphine” and “norbuprenorphine” in the urine as an indicator of urine adulteration [16, 17, 18, 19]. In addition to this high ratio between “buprenorphine” to “norbuprenorphine”, high “naloxone” levels in urine samples may also point to adulteration. Warrington, et al. suggested that urine samples with “naloxone” concentration greater than 2000 ng/mL should raise suspicion of adulteration, likely from dipping buprenorphine/naloxone film into the sample [20]. In their study, “buprenorphine” to “norbuprenorphine” ratios were found to be high along with the elevated “naloxone” levels in the urine samples with suspected adulteration. This supports the notion that “naloxone” concentration in the urine can be used as one of the indicators for urine sample adulteration.
While previous studies utilized urine samples from individuals who were on prescribed buprenorphine/naloxone, the impact of adulteration on substance-free urine and purified water remains unproven. Burns et al. conducted an in vitro study in which naloxone was added to urine samples that came from healthy volunteers who were not on any medications, and they found that “naloxone” was detected in the urine samples [21]. While the elevated “naloxone” level in urine samples may be flagged for adulteration, to our knowledge, there are no studies to date that demonstrate whether high levels of “naloxone” can be detected by dipping buprenorphine/naloxone film in water or urine samples from those who are not taking the combination medication.
The aim of this study is to detect and quantify the presence of “naloxone” in adulterated urine samples from those who are not on buprenorphine/naloxone medication and adulterated purified water samples. The findings of this study should deepen our understanding on the pharmacokinetics and pharmacodynamics of naloxone.
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2. Methods
2.1 Data
This is a urine test experiment study. After the Institutional Review Board (IRB) application was approved at the University of Texas Health at San Antonio (IRB Protocol ID 20220593HU), 12 urine samples from 12 participants were collected at our clinic. These participants were recruited through flyers and word-of-mouth. One of the inclusion criteria stipulates that the participant is not taking any medications or illegal substances such as opioids and marijuana. This criterion was included because certain medications and substances can inhibit or induce relevant CYP450, which might affect naloxone metabolism and consequently the “naloxone” levels in urine [12]. Another inclusion criterion was “healthy” participants without any major health issues. This inclusion criterion was added because naloxone is metabolized in the liver through glucuronide conjugation, and hepatic impairment can alter naloxone metabolism and consequently affect “naloxone” levels in the urine [22]. The last inclusion criterion was the ability to provide >80 mL urine samples. 15 healthy participants who were not on any medications were initially recruited. 3 participants from this initial pool were excluded due to their inability to provide adequate urine volume. Therefore, 12 urine samples from 12 participants were included in this study. At the time of urine collection, these 12 participants were asked to fill out a demographic information form. Their participation to this study was compensated financially for their time and urine sample contribution.
After the 12-urine samples were collected, each sample was then divided into four specimen aliquots with 20 mL each. Then, sublingual generic buprenorphine/naloxone (8 mg/2 mg) film of 12.8 mm width and 22.0 mm length was dipped directly into each urine specimen. In the first aliquot, 1 mm of buprenorphine/naloxone film was vertically dipped into the urine specimen for three seconds (1 mm*3 sec); in the second, half of a film, approximately 11 mm, was vertically dipped for three seconds (half*3 sec); in the third, a full film was dipped for three seconds (full*3 sec); and in the fourth, a full film was dipped for thirty seconds (full*30 sec). That way, we could investigate if area and/or duration of dipping can alter the “naloxone” levels. In addition, four control samples were utilized; (1) room temperature purified water (RT water), (2) water at approximately 97o F (Body Temperature or BT water), (3) 2 mL of urine diluted with 18 mL RT water (10% RT), and (4) 2 mL of urine diluted with BT 18 mL water (10% BT). These four control samples were added to examine if “naloxone” can be detected in water without any human urine and if the temperature and concentration of urine can affect “naloxone” levels in adulterated urine samples.
2.2 Data analysis
All specimens mentioned above were sent to the ARUP laboratory for quantitative tests. The tests included “naloxone” and “creatinine” levels. The results were stored in
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3. Results
The demographic information of the 12 participants was reviewed and is summarized in Table 1.
| Characteristic (N = 12) | Mean ± SD or n (%) | Range |
|---|---|---|
| Age (y) | 32.9 ± 11.3 | 20–57 |
| Sex, Male | 10 (83.3.%) | |
| Ethnicity | 8 (66.7%) | |
| Hispanic | 3 (25%) | |
| White Other | 1 (8.3%) | |
| BMI (kg/m2) | 29.1 ± 5.7 | 21–40.3 |
| Smoking | ||
| Never | 11 (91.7%) | |
| Smoker | 1 (8.3%) | |
| Veterans | 1 (8.3%) | |
| Education | ||
| Some college | 8 (66.7%) | |
| Bachelor’s | 3 (25%) | |
| Others | 1 (8.3%) |
Table 1.
The information is presented as mean ± standard deviation (SD) or mean with
The majority of the participants were male (83.3%), Hispanic (66.7%), nonsmoker (91.7%), and non-veterans (91.7%) with some college education (66.7%). All of them denied any medication use or current medical issues. The average age of the 12 participants was 32.9 years old with a range of 20 to 57 years old. The average BMI was 29.1 kg/m2 with a range of 21 to 40.3 kg/m2.
The “naloxone” and “creatinine” levels were checked, and the average and standard deviation (SD) of the four different specimen aliquots in each sample are listed in Table 2. The “naloxone” level with maximum measurable level was 1000 ng/mL, and any values surpassing it were considered as 1000 ng/mL for the purpose of calculations. Similarly, when the “naloxone” level was lower than the minimal measurable or detectable level (<100 ng/mL), 0 ng/mL was used for the purpose of calculations. The reportable range of “creatinine” was 5-2239 mg/dL, while that of “naloxone” was 100-1000 ng/mL. The “naloxone” was reported with the unit of ng/mL while that of “creatinine” with mg/dL.
| Subjects | Aliquot 1 (1 mm*3 sec) | Aliquot 2 (half*3 sec) | Aliquot 3 (full*3 sec) | Aliquot 4 (full*30 sec) | Creatinine mg/dL |
|---|---|---|---|---|---|
| 1 | <100 | 337 | >1000 | >1000 | 84 |
| 2 | 331 | >1000 | >1000 | >1000 | 165 |
| 3 | >1000 | 110 | 117 | >1000 | 52 |
| 4 | 407 | >1000 | >1000 | >1000 | 27 |
| 5 | 933 | >1000 | >1000 | >1000 | 159 |
| 6 | 226 | >1000 | >1000 | >1000 | 45 |
| 7 | 809 | >1000 | >1000 | >1000 | 97 |
| 8 | 233 | >1000 | >1000 | >1000 | 67 |
| 9 | 524 | >1000 | >1000 | >1000 | 75 |
| 10 | 187 | 969 | 504 | >1000 | 138 |
| 11 | 263 | 601 | 354 | >1000 | 73 |
| 12 | 991 | >1000 | 215 | >1000 | 95 |
| Average | 492.00 | 834.75 | 765.83 | 1000.00 | 89.75 |
| SD | 351.82 | 311.03 | 356.91 | 0.00 | 43.98 |
Table 2.
“Naloxone” levels in 12 urine samples.
Aliquot 1 (1 mm*3 sec) = a buprenorphine/naloxone film was dipped vertically 1 mm for three seconds; Aliquot 2 (half*3 sec) = half film was dipped for three seconds; Aliquot 3 (full*3 sec) = full film was dipped for three seconds; and Aliquot 4 (full*30 sec) = full film was dipped for 30 seconds. SD = Standard Deviation.
Table 2 shows that when buprenorphine/naloxone film was dipped directly into these specimens, “naloxone” was detected in the samples. Although “naloxone” was not detected in one of the 1 mm*3 sec specimens, all of the other specimens had “naloxone” detected in the urine specimens. In particular, all of the specimens with full film dipped for 30 seconds showed high levels of “naloxone” detected with >1000 ng/mL. All the creatinine levels were within normal range (20–400 mg/dL in the ARUP laboratory report) with an average of 89.75 mg/dL.
A statistically significant difference is present when we compared with 1 mm dipped for 3 seconds, half film dipped for 3 seconds, and full film dipped for 3 seconds by a one-way ANOVA (F(2, 33) = [3.401], p = .045). This indicates that the larger the area of film in contact with the urine specimen, the higher the level of “naloxone” detected in the urine. Also, when we compare the specimens that had a full film dipped for 3 seconds (M = 765.83, SD = 356.91) versus 30 seconds (M = 1000, SD = 0) by an independent sample T test, a statistically significant difference of “naloxone” levels was observed; t(22) = −2.273, p = .033). This indicates the longer the time the film was dipped, the higher the “naloxone” levels were detected. Thus, these results indicate that the volume and duration of dipping can influence “naloxone” levels in urine.
Next, the average of “naloxone” levels in the 12 urine specimens, the diluted 10% urine sample, and purified water sample were compared. The results are summarized in Table 3. In the table below, “naloxone”, “creatinine” and “naloxone/creatinine” levels are listed with the units and the cut off levels. Because the unit of “naloxone” was recorded with “ng/mL” while that of “creatinine” was (mg/dL), the ratio between “naloxone” and “creatinine” was listed with (*−4) for an easier understanding.
| Samples | “Naloxone” ng/mL (100–1000) | “Creatinine” mg/dL (5–2239) | “Naloxone”/“Creatinine” (*−4) | |
|---|---|---|---|---|
| Water BT | 1 mm*3 sec | 637 | ND | NA |
| Half*3 sec | >1000 | ND | NA | |
| Full*3 sec | >1000 | ND | NA | |
| Full*30 sec | >1000 | ND | NA | |
| Water RT | 1 mm*3 sec | 347 | ND | NA |
| Half*3 sec | >1000 | ND | NA | |
| Full*3 sec | >1000 | ND | NA | |
| Full*30 sec | >1000 | ND | NA | |
| 10% BT | 1 mm*3 sec | 304 | 7 | 43.43 |
| Half*3 sec | >1000 | 7 | 142.86 | |
| Full*3 sec | 468 | 7 | 66.86 | |
| Full*30 sec | >1000 | 7 | 142.86 | |
| 10% RT | 1 mm*3 sec | ND | 6 | NA |
| Half*3 sec | 761 | 6 | 126.83 | |
| Full*3 sec | 436 | 6 | 72.67 | |
| Full*30 sec | >1000 | 6 | 166.67 | |
| Urine BT | 1 mm*3 sec | 492 | 89.75 | 6.78 |
| (12 urine | Half*3 sec | 834.75 | 89.75 | 11.84 |
| average) | Full*3 sec | 765.83 | 89.75 | 11.26 |
| Full*30 sec | >1000 | 89.75 | 14.4 |
Table 3.
“Naloxone” levels in urine specimens, 10% diluted urine, and purified water.
ND (not detected) indicates that the “naloxone” or “creatinine” was under the detectable level. NA (not applicable) indicates that “naloxone” or “creatinine” was undetected, and thus we were not able to calculate “naloxone”/“creatinine” ratio. BT (body temperature) means that the sample temperature was ~97o F. RT (room temperature) in 10% RT means that the samples were diluted by room temperature purified water. The unit as well as the minimal and maximal measurable levels are listed in the table. “Creatinine” levels of four specimens in one sample are the same because only one urine “creatinine” level in each sample was tested.
Table 3 shows that when the film was dipped into purified water without human urine, “naloxone” was detected. This indicates that “naloxone” detected in urine might be mere dissolvents instead of metabolites. Next, the “naloxone” levels of room temperature water were compared with those of body temperature with one-sample T-test. We found that the “naloxone” levels of room temperature water (M = 836.75, SD = 326.50) were not significantly different from those with body temperature water (M = 909.25, SD = 181.50); t(6) = .388, p = .711). Thus, the temperature difference did not affect “naloxone” levels. Finally, when the “naloxone” levels of body temperature purified water, body temperature 10% urine and body temperature pure urine were compared by a one-way ANOVA, there was no statistical difference (F(2, 9) = [.690], p = .526). Thus, the urine concentration difference did not affect “naloxone” levels. These results indicate that temperature and concentration of urine samples did not affects “naloxone” levels.
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4. Discussion
4.1 Clinical effects of sublingual naloxone
This study demonstrated that high levels of “naloxone” were found in adulterated urine samples from individuals who were not taking any medications. High levels of “naloxone” were also detected in the purified water samples when buprenorphine/naloxone film was dipped into the samples. Thus, these results and those in the previous studies on urine “naloxone” levels suggest that “naloxone” can be detected in adulterated urine samples both from those who are not on buprenorphine/naloxone and those who are on buprenorphine/naloxone [15, 20]. This information can lead to the speculation that “naloxone” detected in urine samples can be mere dissolvents of some fraction of naloxone instead of metabolites, and consequently imply a possibility that “naloxone” in urine samples from the patients taking buprenorphine/naloxone films were the mere dissolvents of some naloxone in the urinary system, considering naloxone’s low bioavailability and extensive first-pass metabolism. Unfortunately, we were unable to distinguish between “naloxone” in urine that has been excreted by nephrons after it was absorbed and metabolized versus some fractions of “naloxone” that have been only dissolved in the system.
The clinical effects of sublingual naloxone on healthy patients have been controversial. For example, Nasser, et al. conducted a research study, in which 43 participants received one single sublingual tablet of brand name Suboxone (buprenorphine/naloxone 2 mg/0.5 mg) and their plasma “naloxone” levels in their blood samples were monitored for up to 168 hours. They found that the participants with severe hepatic impairment had higher and longer “naloxone” levels detected compared with the healthy participants; however, the inactive metabolite, “naloxone 3-β-D-glucuronide” levels were similar between the two groups. They concluded that sublingual buprenorphine/naloxone combined product should be avoided for those who have severe hepatic impairment [4]. This study also suggested that naloxone is metabolized in the liver and may provide clinical effectiveness only for those who have hepatic impairment. The clinical effects of sublingual naloxone on healthy patients remain controversial.
Strickland and Burson reviewed the charts of 561 patients, 11.1% of whom were on buprenorphine/naloxone combination products while the others received mono products. The authors reported that urine “naloxone” was detected in the majority of samples; 97.8% from the 63 patients who were on the dual medication had >1 ng/mL naloxone, and 92.7% of them had 30 ng/mL of naloxone. The authors argued that sublingual naloxone may be absorbed and thus cause unpleasant adverse effects [13].
By contrast, the pharmacological effects of sublingual naloxone have a 10-times lower binding affinity to μ opioid receptors, compared to buprenorphine, namely, rapid opioid receptor dissociation (approximately 6.5 minutes); short half-life of naloxone (60–90 minutes compared to 24–60 hours of buprenorphine); low bioavailability (estimated 3% sublingually); and first-pass bioavailablility, led to the postulation of insignificant clinical effects in sublingual naloxone. After the Food and Drug Administration (FDA) approved buprenorphine/naloxone combination medication in 2002 [24], this combined medication became the standard of care, especially after the Substance Abuse and Mental Health Services Administration (SAMHSA) warned about the misuse potentials of monotherapy [25]. However, the controversy of combined naloxone/buprenorphine versus monotherapy of buprenorphine has yet to be resolved, and further research and discussion on this topic await.
4.2 Adulteration
This study found that the amount of the film that was used to adulterated urine specimen did affect the concentration of “naloxone” detected in the urine. Submerging a small portion of the film for a short period of time (1 mm*3 sec) in this study resulted in an average of 492 ng/mL of “naloxone” levels in the urine samples among the 12 participants. By contrast, the urine samples with full film dipped into the aliquot for 3 seconds (Full*3 sec) yielded an average of 765.83 ng/mL of “naloxone”. These numbers are very similar with those found in the study done by Warrington, et al. on the “naloxone” levels in suspected adulterated urine. They retrospectively reviewed “naloxone” levels of 1223 urine samples from two practice sites and reported that the average “naloxone” level was “633.65ng/mL (range 1-12,161 ng/mL) with 54% of samples <300 ng/mL and 8.0% having >2000ng/mL. One of the sites had increased evidence of urine adulteration and 9.3% of the samples from this site contained > 2000 ng/ml of “naloxone” with an average of 686.8 ng/mL. The other site had no report of urine adulteration and demonstrated an average “naloxone” level of 570.9 ng/mL with 6.4% of samples containing > 2000 ng/mL. This study concluded that extremely high levels of “naloxone” can suggest urine adulteration [20]. Furo also reviewed 97 patient charts with urine drug screening results in an outpatient telemedicine OUD clinic and found that average “naloxone” level was 687.2 ng/mL that ranged from 5 ng/mL to>2000 ng/mL with 15.30% >2000 ng/mL [26].
Heikman, et al. collected 40 urine samples from 32 patients and found much lower average of “naloxone” levels in their study. In their study design, the first group (Group 0: pre-treatment) received parental buprenorphine before buprenorphine/naloxone treatment; the second group (Group 1: stable patients) consisted of stable patients who were on prescribed sublingual buprenorphine/naloxone treatment without any illicit substances in their urine; and the third group (Group 2: unstable patients) had prescribed sublingual buprenorphine/naloxone with unexpected urine test results. The urine samples were collected about 24 hours after the last buprenorphine/naloxone medication dispensation. The study found that the median naloxone level in the stable phase was 60 μg/L (=ng/mL, henceforth ng/mL), ranged from 5 ng/mL to 200 ng/mL while that in the unstable phase was 70 ng/mL, ranged from 70 ng/mL to 1700 ng/mL. They concluded that high “naloxone” level can indicate non-compliance of buprenorphine/naloxone treatment [15].
The average “naloxone” levels in the study by Heikman, et al., [15] was much lower compared with those in the studies mentioned above. This may be because naloxone has a short half-life of 60–90 minutes [22] and because their urine collection was done approximately 24 hours after the last dose of buprenorphine/naloxone medication. The average “naloxone” levels of this current study (e.g. 492 ng/mL of 1 mm*3 sec specimen group and 765.83 ng/mL of full*3 sec group) of the adulterated urine samples from those who were not on buprenorphine/naloxone were similar to those of these previous studies (e.g. 687.2 ng/mL in Furo [26] and 570.9 ng/mL in Warrington, et al. [20]) with the unadulterated urine samples from those were on buprenorphine/naloxone. This similarity should be explored further in relation to the implication of pharmacokinetics and pharmacodynamics of naloxone in the system.
4.3 Clinical applications
The results of this study can be used to help clinicians interpret urine toxicology test results more accurately. In this section, seven simulated cases based on previously encountered results are discussed to present how to apply the findings of this study to the daily practice of OUD treatment. The cases and their associated laboratory values as well as the buprenorphine prescription dosage are summarized in Table 4.
| Cases | Bup (ng/mL) | Norbup (ng/mL) | Bup/Norbup | Nal (ng/mL) | Cre (mg/dL) | Bup /nal prescription (dose per day) |
|---|---|---|---|---|---|---|
| Case 1 | 278 | 635 | 0.44 | 687 | 133.9 | Bup/Nal 16 mg/4 mg |
| Case 2 | >2000 | 17 | 117.65 | >2000 | 97.2 | Bup/Nal 16 mg/4 mg |
| Case 3 | >2000 | >2000 | 1.00 | >2000 | 306.7 | Bup/Nal 16 mg/4 mg |
| Case 4 | >2000 | 10 | 200.00 | <2 (undetectable) | 82.6 | Bup 16 mg |
| Case 5 | >2000 | 17 | 117.65 | 5 | 179.2 | Bup/Nal 16 mg/4 mg |
| Case 6 | >2000 | >2000 | 1.00 | >2000 | 82.5 | Bup/Nal 16 mg/4 mg |
| Case 7 | 286 | 569 | 0.50 | 71 | 67.5 | Bup 16 mg |
Table 4.
7 simulated cases in clinical practice and their associated laboratory values.
Bup = Buprenorphine, Norbup = Norbuprenorphine, Nal = Naloxone, Cre = Creatinine. The units are listed in the top column. The level > 2000 ng/mL was considered 2000 ng/mL for the calculations.
4.3.1 Case 1
[Bup 278 ng/mL, Norbup 635 ng/mL, Bup/Norbup 0.44, Nal 687 ng/mL, Cre 133.9 mg/dL, Bup/Nal 16 mg/4 mg per day, appropriate case].
This is a typical pattern of urine toxicology results. If patients are taking “buprenorphine” daily, “norbuprenorphine” is usually higher than “buprenorphine”, [16] and creatinine is within normal range (20-400 mg/dL) (The ARUP laboratory criteria). Therefore, this is an appropriate urine toxicology case for those who are compliant with buprenorphine/naloxone treatment.
4.3.2 Case 2
[Bup >2000 ng/mL, Norbup 17 ng/mL, Bup/Norbup 117.65, Nal >2000 ng/mL, Cre 97.2 mg/dL, Bup/Nal 16 mg/4 mg per day, inappropriate case].
This is a typical pattern of urine adulteration of having dipped buprenorphine/naloxone film directly into the urine sample. The ratio between buprenorphine/norbuprenorphine is >50 [17]. The very high naloxone level (>2000 ng/mL) confirms this suspicion of adulteration, consistent with the results of this study.
4.3.3 Case 3
[Bup >2000 ng/mL, Norbup >2000 ng/mL, Bup/Norbup 1.0, Nal >2000 ng/mL, Cre 306.7 mg, Bup/Nal 16 mg/4 mg per day, appropriate case].
This case has a high level of naloxone >2000 ng/mL, so there is suspicion of urine adulteration; however, this is an example, in which a high naloxone level does not mean urine adulteration. As Furo’s study found that 15% of urine samples had “naloxone” levels >2000 ng/mL [26], a high level of naloxone can also be due to other reasons. This case illustrates that we can use a high “naloxone” level to confirm urine adulteration only if the buprenorphine/naloxone ratio is >50:1; “naloxone” level itself cannot be the main determining factor of urine adulteration. In other words, the judgment of urine adulteration should be based on high buprenorphine/norbuprenorphine ratio. Therefore, this is an appropriate case of buprenorphine/naloxone treatment. One might wonder why is this patient’s “naloxone” level so high? The answer is that this patient might be dehydrated at the time of this urine collection, indicated by the high creatinine level, so the buprenorphine, norbuprenorphine, naloxone and creatinine are all consequently very high. If this patient were well hydrated, the results should have been similar to Case 1 with a much lower creatinine level. Thus, this example should be appropriate for the buprenorphine/norbuprenorphine treatment.
4.3.4 Case 4
[Bup >2000 ng/mL, Norbup 10 ng/mL, Bup/Norbup 200, Nal <2 ng/mL, Cre 82.6 mg/dL, Bup 16 mg per day, inappropriate case].
This case is an adulteration case in which the patient is prescribed buprenorphine only medication. This patient has not taken this medication for a while, indicated by the low norbuprenorphine level. This case has a high ratio of buprenorphine/norbuprenorphine, but naloxone is undetectable, which means that prior to the urine collection, buprenorphine medication has been crushed into powder and dissolved into the sample. As a result, the ratio between buprenorphine/norbuprenorphine is high (>50), but there is undetectable naloxone level (<2 ng/mL). Undetectable naloxone level should be always suspected with buprenorphine monotherapy.
4.3.5 Case 5
[Bup >2000 ng/mL, Norbup 17 ng/mL, Bup/Norbup 117.65, Nal 5 ng/mL, Cre 179.2 mg/dL, Bup/Nal 16 mg/4 mg per day, inappropriate case].
This is another case of urine adulteration as indicated with a high buprenorphine/norbuprenorphine ratio (>50:1). However, “naloxone” is detected at a very small amount. Because of the high buprenorphine/norbuprenorphine ratio, we expect a high level of “naloxone” if buprenorphine/naloxone was dipped in this urine sample. Otherwise, we expect undetectable level of “naloxone” if buprenorphine only medication was dissolved into the urine sample. The small amount of “naloxone’s being detected means that this patient probably has taken buprenorphine/naloxone at least 24 hours before this urine collection [15], and thus there was a small amount of “naloxone” residue detected. Compared with buprenorphine, the naloxone’s half-life is much shorter, and thus, a low level of naloxone can be detected while relatively higher level of buprenorphine and norbuprenorphine still remained in the system after an intermittent use of buprenorphine/naloxone. In addition, this patient tampered the urine sample with buprenorphine only medication, which is indicated by the high buprenorphine/norbuprenorphine ratio. If buprenorphine/naloxone combined medication was dipped into this urine sample, the naloxone level would have been much higher as indicated by the results of this study, unless the patient has some issues of naloxone metabolism such as the enzyme to metabolize naloxone is inhibited by certain medication(s) or genetically. This patient is prescribed with buprenorphine/naloxone combined medication, so it is uncertain as to why this patient added additional buprenorphine to the urine.
4.3.6 Case 6
[Bup >2000 ng/mL, Norbup 2000 ng/mL, Bup/Norbup 1.0, Nal 2000 ng/mL, Cre 82.5 mg/dL, Bup/Nal 16 mg/4 mg per day, appropriate case].
This case has high “buprenorphine”, “norbuprenorphine” and “naloxone” levels, so we can speculate dehydration; however, “creatinine” level is not as high as Case 3, and therefore, we can rule out dehydration. The ratio between buprenorphine to norbuprenorphine is <50:1, so it is unlikely that this sample is adulterated. This might be a case with high metabolism patient, which causes high levels of all components. This is rare but can happen. Thus, this is another example of a high naloxone level, but it does not mean urine adulteration.
4.3.7 Case 7
[Bup 286 ng/mL, Norbup 569 ng/mL, Bup/Norbup o.5, Nal 71 ng/mL, Cre 67.5 mg/dL, Bup 16 mg per day due to naloxone allergies, inappropriate case].
This patient has an appropriate “buprenorphine” to “norbuprenorphine” ratio (<50:1) with low level of “naloxone”; however, this result is not appropriate because this patient was on buprenorphine monotherapy due to naloxone allergies. The detected “naloxone” suggests that she has been taking buprenorphine/naloxone despite her being prescribed buprenorphine monotherapy. Buprenorphine only product has a higher street value than buprenorphine/norbuprenorphine combined medication by approximately 20% [27]. Therefore, some patients might trade their buprenorphine with buprenorphine/naloxone combination medication for the marginal profit.
In summary, as this study found, a high concentration of “naloxone” can confirm suspected urine adulteration if buprenorphine/norbuprenorphine ratio is >50:1. However, a high level of “naloxone” alone does not necessarily involve urine adulteration, especially without a high ratio of buprenorphine/norbuprenorphine. The patient might have dehydrated at the time of urine collection, or the patient might be a fast metabolizer of buprenorphine/naloxone. Therefore, we should monitor creatinine level because it can indicate the hydration status of the patient [28], as dehydration can cause high levels of “naloxone” in addition to high “buprenorphine” and “norbuprenorphine” levels. Furthermore, a patient with high metabolism can cause high levels of “buprenorphine”, “norbuprenorphine” and “naloxone” without a corresponding high “creatinine” level. Finally, if “naloxone” is observed in urine sample, we can suspect that the patient is taking or had taken buprenorphine/naloxone combined medication, while if no naloxone is identified, the patient is on buprenorphine monotherapy.
4.4 Clinical implications
The results of this study have implications on clinical practice in the care of OUD patients. With the increased use of unobserved urine toxicology collection in practice [29], the appropriate interpretation of urine drug tests remains a challenging but vital component of comprehensive patient care. Results from this study may suggest that the “naloxone” detected in adulterated urine samples may be dissolved “naloxone” instead of metabolized naloxone, or a fraction of naloxone at least. There are also healthcare policy implications with the controversy of mono-product formulation use in the United States as a harm reduction approach in the treatment of OUD [9]. Buprenorphine mono-product is used widely in other areas of the world [30] and has been suggested as a medication treatment option for patients struggling with adverse effects, thought to be potentially from naloxone absorption from sublingual buprenorphine/naloxone combined medication [9]. With increasing research findings that raise the concern of using “naloxone” levels in urine as an indicator of absorption of naloxone and potentially the cause of patient-reported adverse effects, these findings reinforce the recommendation for clinicians to also consider possible urine adulteration, resulting in elevated “naloxone” levels.
A high level of “naloxone” itself should trigger further assessment of the patient buprenorphine/naloxone regimen; however, by itself there is insufficient evidence to guide care. This information should be synthesized with other available urine toxicology screen parameters (buprenorphine, norbuprenorphine, creatinine and buprenorphine/norbuprenorphine ratio) to better inform the clinical picture. Moreover, the continuation of buprenorphine as a medication for OUD yields more benefits than risks in the public health approach of addressing opioid overdose mortality. Urine toxicology is only one piece of information to be used in a clinical evaluation. The overall goal of improvement of multiple biopsychosocial domains and patient-centered outcomes remains the driving force in clinical decision-making.
Finally, the results of this study might support the stipulation that sublingual naloxone can be minimally absorbed and metabolized in the system and thus exerts insignificant effects clinically when taken sublingually, which might consequently contribute to the controversy on buprenorphine/naloxone combined product versus buprenorphine monotherapy in the buprenorphine induction process [31, 32]. The results of this study would give us an insight on these controversial issues.
4.5 Limitations
The limitations of this study include the small sample size and reliance on subjects’ report of substance and medication use. Findings from this study may be used to apply for funding to support a larger sample size study and additional substance testing of all samples. While the potential effects of unreported substance use are unknown, the overall trend in this study’s results is unlikely to be impacted. An additional limitation is that this study evaluated the addition of varying buprenorphine/naloxone levels via film into water and urine samples as a proxy for adulteration. The actual adulteration techniques used by individuals may vary widely. A clinical trial evaluating “naloxone” levels in adulterated and non-adulterated urine samples of patients prescribed buprenorphine/naloxone would be difficult in nature. To address the range in adulteration techniques, we used multiple categories of surface areas, dipping duration, temperatures, and concentrations of both urine and water samples to assimilate a variety of methods and observe the resulting “naloxone” levels. Finally, we also acknowledge that some clinicians use urine toxicology that do not report “naloxone” levels in certain clinic practices, and therefore these findings may not be clinically useful to all providers.
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5. Conclusions
The urine “naloxone” level by dissolving buprenorphine/naloxone film is very sensitive to the area and the time that the film come into contact with the urine samples. Attempted adulteration of the urine sample is likely yield supratherapeutic levels of “naloxone”. At this time, there is no consensus as to what level to set the “naloxone” concentration as a parameter to determine the legitimacy of patient urine samples except some research reports. While “naloxone” level is a simple component to test for in resource-challenged practices, other metabolites such as “buprenorphine” and its metabolite levels should be examined to guide clinical decisions. In summary, the results of this study have provided a further insight into interpreting urine drug screening test results for OUD patients with buprenorphine treatment. Strict monitoring of urine toxicology is by no means a punitive process but to improve the outcome of OUD treatment. Future study can focus on differentiating naloxone molecules that were dissolved “naloxone” versus those that were renally excreted, which can enhance our understanding on the pharmacokinetics and pharmacodynamics of naloxone.
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Acknowledgments
We would like to express our sincere and deep appreciation for the editors and reviewers who provided us with valuable feedback. We would also thank Editage for editing the final version of this manuscript.
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Authors’ contributions
H.F. carried out the research experiment and wrote the first draft of the paper excerpt discussion and conclusion sections. T.H. wrote the discussion and conclusion sections and completed the manuscript with support with S.A. who edited the draft form of this manuscript. Y.Z. completed the manuscript by editing the final version.
Note
Correspondence concerning this article should be addressed to Hiroko Furo (
Ethics approval and consent to participate
The Institutional Review Board (IRB) application to the University of Texas Health at San Antonio was approved (IRB Protocol ID: 20220593HU), and the written consents were obtained by all participants.
Availability of data and materials
The datasets produced for the data analysis for this study are not publicly available due to the confidentiality of participants but are available in a de-identified form from the corresponding author on request.
Abbreviations
| “Naloxone” levels in 12 urine samples—1 mm*3 sec | buprenorphine/naloxone film was dipped vertically 1 mm for three seconds |
| Half*3 sec | half film was dipped for three seconds |
| Full*3 sec | full film was dipped for three seconds |
| Full*30 sec | full film was dipped for 30 seconds |
| RT | room temperature |
| BT | body temperature ~ 97oF |
| ND | not detected |
| SD | standard deviation |
| N | number |
| BMI | body mass index |
| M | mean |
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Notes
- In order to differentiate "naloxone" found in urine versus other naloxone format such as naloxone in buprenorphine/naloxone combined medication, "naloxone" and other components found in specimens are expressed with quotation marks around them as "naloxone."